Isolates of Leptospira spp. were cultured from water sources at five different sites in central Iowa in the Midwestern United States and characterized by whole-genome sequencing. Isolates were helix-shaped and motile. Genome sequence analyses determined that the isolates could be clearly distinguished from other species described in the genus Leptospira and included one species that belonged to the pathogen subclade P1, one species that belonged to the pathogen subclade P2 and three species that belonged to the saprophyte subclade S1. The names Leptospira gorisiae sp. nov. (type strain WS92.C1^T=NVSL-WS92.C1^T=KIT0303^T), Leptospira cinconiae sp. nov. (type strain WS58.C1^T=NVSL-WS58.C1^T=KIT0304^T), Leptospira mgodei sp. nov. (type strain WS4.C2^T=NVSL.WS4.C2^T=KIT0305^T), Leptospira iowaensis sp. nov. (type strain WS39.C2^T=NVSL-WS39.C2^T=KIT0306^T) and Leptospira milleri sp. nov. (type strain WS60.C2^T=NVSL-WS60.C2^T=KIT0307^T) are proposed.

Staphylococcus pseudintermedius is the most common opportunistic pathogen in dogs and methicillin resistance (MRSP) has been identified as an emerging problem in canine pyoderma. Here, we evaluated the antimicrobial resistance (AMR) features and phylogeny of S. pseudintermedius isolated from canine pyoderma cases in Argentina (n = 29) and the United States (n = 29). 62% of isolates showed multi-drug resistance. The AMR genes found: mecA, blaZ, ermB, dfrG, catA, tetM, aac(6′)-aph(2″), in addition to tetK and lnuA (only found in U.S. isolates). Two point mutations were detected: grlA(S80I)-gyrA(S84L), and grlA(D84N)-gyrA(S84L) in one U.S. isolate. A mutation in rpoB (H481N) was found in two isolates from Argentina. SCCmec type III, SCCmec type V, ΨSCCmec₅₇₃₉₅ were identified in the Argentinian isolates; and SCCmec type III, SCCmec type IVg, SCCmec type V, and SCCmec type VII variant in the U.S. cohort. Sequence type (ST) ST71 belonging to a dominant clone was found in isolates from both countries, and ST45 only in Argentinian isolates. This is the first study to comparatively analyze the population structure of canine pyoderma-associated S. pseudintermedius isolates in Argentina and in the U.S. It is important to maintain surveillance on S. pseudintermedius populations to monitor AMR and gain further understanding of its evolution and dissemination.

Taylorella equigenitalis , the causative agent of contagious equine metritis (CEM), is endemic in several countries throughout the world and impacts the global movement of horses. It presents with symptoms ranging from non-clinical to endometritis, vaginitis, and acute infertility in females, while males remain asymptomatic. Studies of the disease have focused primarily on understanding pathology, transmission dynamics, diagnostics, and phenotypic and genotypic characteristics. The goal of this study was to establish a more comprehensive global phylogenetic structure and generate reference-quality genomes for each major clade of the tree where such data were absent. Seven of these genomes along with 44 other publicly available genome assemblies, including Taylorella asinigenitalis , were then used to predict genomic islands. Exploration of the NCBI annotations in the genomic islands indicate links to virulence and ecological fitness. An additional 97 T. equigenitalis and 48 T. asinigenitalis short read sequenced isolates from this study and 215 publicly available genomes were used to examine distribution of each genomic island across the phylogenetic tree for evidence of horizontal movement. The stability of these genomic islands in their natural environment was evaluated utilizing 141 isolates recovered from a recent CEM outbreak in the United States. The results obtained in this study demonstrate mobile genomic islands in T. equigenitalis genomes, sites of concern for their contribution to the potential virulence of the organism.

Background: Mannheimia haemolytica is a gram-negative bacterium that typically resides in the upper respiratory tract of healthy cattle but can invade the lower respiratory tract in immunocompromised cattle, contributing to bovine respiratory disease (BRD). The rise of antimicrobial resistance (AMR) in M. haemolytica strains, especially those linked to BRD, is a growing concern. This study aimed to investigate AMR phenotypes and genotypes in M. haemolytica isolates collected during the NAHLN AMR Pilot Project (2018–2022) and assess the use of a previously developed single nucleotide polymorphism genotyping scheme to classify strains and identify associations between genotypes, AMR genes, quinolone point mutations (QPMs) and mobile genetic elements in diseased cattle. Results: Of 2743 isolates tested for antimicrobial susceptibility, 66.7% (1830/2743) were pan-susceptible, while 33.3% (913/2743) showed resistance to at least one antimicrobial, primarily tetracyclines. Among the 848 sequenced isolates, 36.7% (311/848) harbored AMR genes and/or QPMs. Of these, 59.8% (186/311) were phenotypically classified as multidrug-resistant; commonly detected AMR genes included aadA , ant(2’’)-Ia , bla _OXA−2 , mph (E), msr (E), and tet (H). The study found that 88.3% (749/848) of isolates belonged to genotype 2, a group previously associated with BRD and AMR. Notably, 95.8% (298/311) of isolates harboring AMR genes were from genotype 2. Although most clinical isolates of M. haemolytica were pan-susceptible and lacked AMR genes, a substantial proportion of isolates exhibited AMR, with a high degree of concordance between genotype and phenotype across all antimicrobials tested. Most clinical isolates (749/848, 88.3%), primarily obtained from the cattle lungs rather than the upper respiratory tract, were classified as genotype 2. These isolates harbored a T4SS integrative and conjugative element (ICE, 283/848, 33.4%) as well as an integrative and mobilizable element (IME, 59/848, 7.0%) containing AMR genes. Conclusions: This study highlights a strong association between genotype 2 M. haemolytica and AMR, with a significant prevalence of multidrug-resistant strains. The findings underscore the importance of genotypic characterization and whole-genome sequencing (WGS) for monitoring AMR in M. haemolytica and informing future management strategies for BRD.

Background: Genetic stability of live-attenuated infectious laryngotracheitis virus (ILTV) vaccines is essential for consistent efficacy and safety; however, marker-based assessments targeting partial genes are often insufficient given the virus’s large, structurally complex genome. The ILTV genome contains long internal inverted repeats (IRs) that can give rise to genomic isomers, complicating short-read assembly and accurate resolution of genome structure. Methods: To overcome these limitations, we used a hybrid whole-genome sequencing (WGS) strategy, combining Oxford Nanopore Technologies (ONT) long reads to improve assembly contiguity with Illumina short reads for high-accuracy polishing at the single-nucleotide level. Using this approach, we generated complete de novo genome assemblies for the commercial Serva and Salsbury #146 vaccine strains. Results: The assemblies showed high sequence concordance with targeted regions validated by Sanger sequencing. Whole-genome analysis further enabled detection and independent validation of a structural inversion in the unique short (US) region of the Salsbury strain, consistent with herpesvirus genome isomerization. To enable phylogenetic inference despite structural variability, we performed a pangenome-based analysis to define a conserved core-genome dataset that robustly resolved vaccine-associated lineages, separating Serva- and Salsbury-derived strains. Conclusions: Collectively, these findings show that a hybrid WGS workflow can generate high-confidence genome assemblies for the specific commercial ILTV vaccine vials analyzed and can support QC-relevant detection of major structural variations. Because this study is cross-sectional (two strains; single lot/vial per strain), it cannot distinguish potential biological lot-to-lot variation from methodological differences, and a comprehensive genetic stability evaluation will require applying this workflow across defined passage levels and/or multiple production lots.